Project Summary: Melanoma is the most lethal skin cancer with over 90,000 new cases in United States each year. The risk of developing melanoma is substantially greater for people with lightly pigmented skin (1:38 in the US) than those with darkly pigmented skin (1:1000 in the US). Although this discrepancy is generally attributed to the physical UV shielding effect of melanin pigment, the sun protective factor (SPF) of melanin is only 2-3, which seems insufficient to explain the 40-fold difference of melanoma incidence in darkly and lightly pigmented skin. Our preliminary data show that primary human melanocytes (MCs) from lightly pigmented skin produce less melanin relative to primary MCs from dark skin, and this is associated with a decreased MC differentiation state and increased proliferative capacity. Additionally, the proliferation and differentiation differences between light and dark MCs persist upon transformation of primary MCs with medically relevant melanoma drivers (BRAFV600E, CDK4R24C, P53R248W, and hTERT). To test if these differences translated into different melanoma phenotypes in vivo, I engineered architecturally faithful 3-D human skin and orthotopically xenografted the tissue on SCID mice. My preliminary data show that transformed light MCs form early melanomas, whereas transformed dark MCs do not. This observation led us to question whether this stark difference was mediated by melanin synthesis itself, or by intermediates of melanin synthesis, such as dihydroxyphenylalanine (DOPA). We found that darkly pigmented MCs contain approximately 300% more DOPA, as compared to light MCs. Previous work along with my preliminary data show that DOPA serves as a signaling molecule and inhibits proliferation. Specifically, I found that exogenously supplied DOPA induces melanin synthesis in light MCs and melanoma cells, but has no effect on primary dark MC, which likely contain saturating levels of endogenously produced DOPA. The mechanism(s) of the DOPA anti-proliferative effect are currently unknown, but my preliminary data show that DOPA inhibits canonical Gq GPCR pathways leading to a decrease in MAPK and PI3K/AKT signaling pathways. In a collaborative effort with Dr. Bryan Roth at the University of North Carolina, we conducted a functional screen to test whether DOPA binds to any of the ~320 non-olfactory GPCRs in the human genome. The top hit was the Gq-coupled metabotropic glutamate receptor 5 (GRM5), which is a known melanoma driver. Aim 1 will focus on determining the mechanism of DOPA?s anti-proliferative effect, by specifically focusing on its inhibitory role of GRM5, or other top hits from our GPCR screen. Additionally, I determined that DOPA inhibits growth of medically relevant syngeneic BRaf-driven melanoma in vivo. Aim 2 seeks to further understand the endogenous and exogenous roles of DOPA in melanoma inhibition in medically-relevant melanoma models in vivo. Together, these aims will help to define the underexplored melanoma pathobiology responsible for increased melanoma in lightly pigmented people, and may identify new therapeutic targets to benefit melanoma patients.